Reports estimate that there are over 17 million individuals with diagnosed Type 2 Diabetes (T2DM) in the US, and an additional ~ 6 million undiagnosed individuals unaware of having (or at risk of having) T2DM. This number of afflicted individuals is expected to double in the US in the next 40 years. Current costs for diabetes in the US are estimated at $174 billion (~$1 of every $10 health care dollars), with cardiovascular disease (CVD) - the major cause of morbidity and mortality for individuals with diabetes - being the largest contributor to direct and indirect costs. Currently, there are two common biomarkers used in the detection and monitoring of T2DM: blood glucose and glucose modified hemoglobin. In preliminary studies, we have built upon these current diagnostic tests by viewing subtle, dynamic changes in the proteome of patients with T2DM and CVD. Briefly, molecular differences were observed in a number of plasma proteins, which during univariate analysis resulted in good separation between healthy and T2DM cohorts. In subsequent treatments, data were grouped into panels and correlated to a three-dimensional view of glycation vs. oxidative-stress vs. perturbations in the insulin-signaling pathway. Scatter plots revealed subtypes in the T2DM population exhibiting differential glycation/oxidative stress/signaling relationships, in particular low glycation, but high oxidative stress - a sub-clinical driver of CVD. These studies lead to creating a multidimensional view of the T2DM/CVD continuum aligned with the pathobiologies of the diseases. Here, we will continue our investigations using > 700 patients in total, across a continuum of healthy - to - pre-T2DM - to - T2DM - to negative CVD outcome. We will follow a systematic biomarker development pathway to: (Specific Aim 1) identify additional candidate biomarkers for detection and monitoring T2DM and CVD; (Specific Aim 2), verify the utility of the candidate biomarkers using larger populations; which provides the knowledge to (Specific Aim 3) design and implement 2nd-generation assays that group the T2DM and CVD comorbidity protein biomarkers into multiplexed assays. The overall goal of this proposed research is to produce verified protein biomarkers along with accompanying assays and methods of data evaluation that will result in the improved monitoring and treatment of type 2 diabetes mellitus and cardiovascular disease comorbidities.